The influence of conserved tyrosine 30 and tissue-dependent differences in sequence on ferritin function: use of blue and purple Fe(III) species as reporters of ferroxidation

摘要: Ferritins uniquely direct the vectorial transfer of hydrated Fe(II)/Fe(III) ions to a condensed ferric phase in central cavity soluble protein. Secondary, tertiary and quaternary structure are conserved ferritin, but only five amino acid residues among all known ferritins. The sensitivity ferroxidation rates small differences primary sequence between ferritin subunits that cell-specifically expressed or conservative replacement tyrosine 30 residue was demonstrated by examining recombinant (frog) H-type (red blood cell predominant) M-type subunit (liver proteins which both fast ferritins; form two differently colored Fe(III)-protein complexes absorbing at 550 nm 650 nm, respectively. convenient reporters interaction because they transient contrast Fe(III)-oxy measured past 310–420 nm, stable contributions from mineral itself. A650-nm species formed 18-fold faster M-subunit protein than did 550-nm H-subunit even though ferroxidase same; Vmax fivefold Hill coefficents were identical (1.6), suggesting similar mechanisms. In substitution phenylalanine for (located core four-helix bundle) slowed tenfold, whereas changing surface 25 28 had no effect. Fe(III)-tyrosinate fortunately not changed mutation, based on resonance Raman spectrum, remained suitable reporter interactions. Thus, A550/650 nm can also report post-oxidation events such as transport through impact Y30F formation combined with Mossbauer spectroscopic studies showed parallel multiple Fe(III) postoxidation (three dinuclear oxy one trinuclear species) (A. S. Periera et al., Biochemistry 36 : 7917–7927, 1997) loss several multimeric alteration human H-ferritin (E. R. Bauminger Biochem J. 296 : 709–719, 1993), indicate least pathways Fe oxidation/transfer is center bundle influenced structural features dependent 30.